The goal of this project is to evaluate novel candidate protein biomarkers for Alzheimer's disease that we have previously identified in CSF using a non-biased proteomics approach (two-dimensional difference in gel electrophoresis [2D-DIGE] coupled with liquid chromatography and tandem mass spectrometry [LCMS/MS]) that compared the CSF proteomes of samples derived from two groups of well-characterized subjects: one with mild dementia of the Alzheimer's type (DAT) and evidence of amyloid deposition in the brain (low CSF Ab42 levels) (N=24);and another without demenia and no evidence of amyloid deposition in the brain (high CSF Ab42 levels) (N=24). To evaluate these candidate biomarkers, in Aim 1 we will develop specific quantitative assays (e.g. ELISA) and validate them assays using CSF from the original 'discovery'cohort of subject samples. In Aim 2, we will apply these validated assays to a Iarger, independent set of CSF samples obtained from well-characterized volunteer subjects followed longitudinally at the WU ADRC, in order to evaluate each assay's diagnostic and prognostic utility. In Aim 3, we propose to perform 'top down'proteomics on a small number of these candidate biomarkers that exhibited aberrant patterns of migration on 2D-DIGE suggestive of unusual post-translational modifications characterizing such distinguishing modifications will enable the development of additional biomarker assays in the future, and may yield insights into the pathophysiology of AD.